Hepatitis C virus (hereinafter referred to as “HCV”) is a major causative virus of post-transfusion non-A, non-B hepatitis. A cDNA of its gene was cloned in 1989. To date, many studies have been conducted on HCV using cloned gene cDNAs. In particular, socially important results such as prevention of infection and establishment of diagnostic methods have been achieved. Thus, the incidence of post-transfusion HCV infection is almost eliminated at the present. However, the number of HCV-infected patients is estimated to account for several percentages of the world's total population.
The hepatitis caused by HCV infection tends to be chronic and persistent. This leads to chronic hepatitis, which is known to develop into cirrhosis and then liver cancer at a very high rate. Thus, reliable treatment of hepatitis after HCV infection is an essential task.
Interferon (IFN) therapy is generally performed as a method for treating chronic hepatitis C. However, there are problems with IFN therapy, such as only 30% efficacy rate, frequent induction of adverse effects including fever, and high drug prices. Studies have been conducted to assess the types of IFN, administration method and dose. Furthermore, the efficacy rate is expected to improve as a result of the development of consensus IFNs. Also, therapies that use a combination of an IFN with an antiviral agent such as Ribavirin are under trial. However, to date, such therapeutic methods have not become reliable.
On the other hand, recently, a method for suppressing target gene expression using a double-stranded RNA against a target gene was developed as a method for suppressing the expression of a specific gene in animal cells in vivo (Non-patent Document 1). This method is called “RNA interference (RNAi)”, which is a phenomenon in which a double-stranded RNA (dsRNA) introduced into cells causes specific degradation of a cellular mRNA corresponding to the dsRNA sequence, and the expression of a protein encoded by the mRNA is blocked. RNAi is an effective method for assessing the function of a novel gene by inhibiting its gene expression, and it is widely used in functional analyses of genes of C. elegans, Drosophila, etc.
The internal ribosomal entry site (IRES) which contains a 5′ untranslated region and a portion of the core region is known to play an important role in the translational initiation and protein synthesis of HCV (Non-patent Document 2). The IRES region which plays an important role in HCV replication has various higher-order structures such as a stem region which forms a stem loop. There are many reports on the 5′ untranslated region, IRES, and stem region of HCV (Non-patent Documents 2 to 12). As described above, the IRES region is a gene region important for HCV replication, and its primary structure (nucleotide sequence) is well conserved among HCVs having different genotypes.
There are multiple HCVs with different genotypes. Such HCVs include, for example, HCJ6, HCJ8, HCV-1, HCV-BK, HCV-J, HCVSHIMO, JCH1, JCH3, JFH1, R24, R6, and S14J. An IRES region that exhibits a higher identity among the sequences of multiple HCVs with different genotypes is preferably targeted to cover the HCV RNAs of multiple HCVs with different genotypes. However, the conformation of the IRES region is complex because the region exerts its translational initiation function by its higher-order structure. Thus, it has been difficult to identify siRNA sequences that exhibit highly efficient RNAi activity using a conventional algorithm for identifying siRNA sequences.
The present inventors have identified and reported siE sequences that exhibit a highly efficient RNAi activity (Non-patent Document 13). However, there has been a need to identify siRNA sequences that have a more effective RNAi activity against hepatitis C virus RNAs.